Search Results (159)

This work reports the synthesis and characterization of a new molecular hybrid 4, created by combining 1,4-naphthoquinone with the drug zidovudine (AZT) through an azide-alkyne cycloaddition reaction catalyzed by Cu¹⁺. In vitro studies assessed the anti-trypanosomatid activity of hybrid 4, along with its precursors and synthetic intermediates (1, 2, and 3), against Trypanosoma cruzi (T. cruzi Tulahuen C2C4 LacZ), Trypanosoma brucei (T. b. brucei 427), and Leishmania infantum, as well as cytotoxicity in RAW 264.7 macrophages and LLC-MK2 cells. The biological results confirm the molecular design, showing that the new hybrid is effective against both epimastigotes and amastigotes of T. cruzi (IC₅₀ = 22.26 ± 5.78 μM and 143.10 ± 5.79 μM, respectively), with approximately 4.5-fold better capacity than AZT to inhibit the epimastigote form. Additionally, the hybrid was also active against bloodstream T. b. brucei (IC₅₀ = 54.47 ± 6.70 μM), with approximately 2.2-fold better capacity than AZT to inhibit this parasite. It also shows low toxicity in RAW 264.7 macrophages (CC₅₀ > 200 μM) and LLC-MK2 cells (CC₅₀ > 200 μM). For example, hybrid 4 exhibited approximately a 6.6-fold higher SI than 1,4-naphthoquinone 1 against T. cruzi amastigotes. In this context, the work contributes to the broader knowledge base guiding the design of hybrid molecules for antiparasitic chemotherapy. It provides a rational foundation for preparing subsequent, more potent analogues. Full article

Trypanosoma cruzi is the protozoan parasite responsible for Chagas disease, a neglected tropical disease caused by trypanosomatids. Its success as pathogen relies on remarkable metabolic adaptability, stress tolerance, and complex interactions with mammalian hosts. Among the proteins contributing to these processes, nucleoside diphosphate kinases (NDPKs) and arginine kinase (AK) have emerged as central enzymes for parasite metabolism. NDPKs, beyond their canonical role in nucleotide homeostasis, are implicated in DNA repair and oxidative stress responses and are also secreted enzymes. AK, on the other hand, serves as a unique energy-buffering system absent in mammals, supporting parasite growth and adaptation to oxidative and metabolic stresses, including modulation of host immunity. Both enzymes display distinct subcellular localizations all along the parasite and through the life cycle, linking them to multiple roles important for parasite biology and survival. Recent studies have highlighted the impact of interfering these enzymes with several compounds on the viability of the organisms, suggesting new avenues to explore them as drug targets. This review provides a general overview of NDPKs and AK in T. cruzi, aiming to underline their relevance to a broader context of trypanosomatids. Their study not only broadens our understanding of parasite biology but also opens perspectives for applied research, including therapeutic alternatives for Chagas and related diseases. Full article

Zoological gardens represent unique sites for vector and vector-borne disease studies. They offer suitable breeding habitats for vector development and a diverse range of vertebrate hosts for blood feeding of insect vectors. This study aimed to assess the prevalence of avian blood parasites (Haemosporida, Trypanosomatida) in wild-caught mosquitoes (Culicidae) and Culicoides biting midges (Ceratopogonidae) from the largest and oldest zoo in Lithuania. Insects were collected in May–August 2023 using UV-light, CDC and BG-Sentinel traps; collected material was analysed using both microscopy and PCR-based methods for parasite detection. Overall, 504 parous biting midges (10 species) and 59 mosquitoes (three species) were investigated. Haemosporidians (Haemoproteus minutus (hTURDUS2), H. homogeneae (hSYAT16), and H. asymmetricus (hTUPHI01)) were identified in 5.4% of the 174 tested biting midges. Haemoproteus asymmetricus hTUPHI01 sporozoites were seen in only one individual of Culicoides kibunensis. Of 108 Culicoides females, 3.7% carried trypanosomatids—parasites infecting birds (Trypanosoma bennetti group) and mammals (T. theileri group). Among the 59 tested mosquitoes, two (3.4%) Cx. pipiens/torrentium mosquitoes were found to be PCR-positive for trypanosomatids (T. culicavium and Crithidia brevicula). No haemosporidian parasite DNA was detected in the mosquitoes examined. This pilot study indicates that avian blood parasites circulate within the Lithuanian Zoo, highlighting the need for further research on transmission pathways, vector–host interactions, and potential risks. Full article

Background: In recent years, compounds known as Proteolysis Targeted Chimeras (PROTACs) have revitalized the field of bioactive molecule design. These compounds promote proteolysis of therapeutic targets by recruiting them to ubiquitin ligases. One of the most commonly used classes of compounds in the synthesis of PROTACs are immunomodulatory imides (IMiDs), such as thalidomide (TLD), which interact with the E3 ligase CRL4^CRBN via the CULT domain of the cereblon protein (CRBN). This domain has been identified in proteins across various phylogenetic groups, including trypanosomatids, leading to the hypothesis that IMiD-derived PROTACs should be active in these organisms. Methods: The trypanocidal activity of the PROTAC dBET1 and its separated components (JQ1 and TLD) were assayed using a T. cruzi strain expressing β-glalactosidase. Potential CRL4-E3L complexes from humans and trypanosomatids were assembled in silico with MultimerMapper. The IMiD-binding site of HsCRBN and its trypanosomatid homologs were analyzed using molecular dynamics and docking simulations. Results: We demonstrate that the compound dBET1 does not function as a PROTAC in Trypanosoma cruzi. In silico structural analysis of CRL4-E3L complex orthologs revealed that the trypanosomal CULT-containing protein is not part of such a complex. Molecular dynamics simulations showed that the pocket of this CULT domain is smaller than that of mammalian CRBN and cannot accommodate IMiDs within. Conclusions: We underscore the importance of functional and structural validation in drug discovery, particularly when extrapolating mechanisms between evolutionarily distant species. While PROTACs hold promise in human therapeutics, our work advocates for re-evaluating the rationale behind thalidomide-based PROTACs in trypanosomatid research. Full article

Background/Objectives: Chagas disease is a neglected tropical disease caused by the protozoan parasite Trypanosoma cruzi, which currently affects around 8 million people worldwide. The therapeutic arsenal against T. cruzi is so far limited to only two approved drugs, benznidazole and nifurtimox, that have considerable side effects and limited efficacy in the chronic stage of the disease. Here, we have resorted to supervised phenotypic machine learning models to explore drug repurposing opportunities and identify potential new therapeutic solutions for Chagas disease. Methods: More than 100,000 bioactivity data points were retrieved from ChEMBL and carefully curated according to the data-centric machine learning paradigm. After curation, two datasets comprising 344 compounds tested against T. cruzi Y strain trypomastigotes and 785 compounds tested against Tulahuen strain amastigotes were obtained and used to infer ensemble learning models with excellent average and early enrichment metrics in retrospective screening experiments (AUROC > 0.96 and EF_0.01 > 58). A prospective screening campaign was then performed on DrugBank and the Drug Repurposing Hub databases, submitting eight in silico hits for experimental confirmation. Results: Six of the in silico hits confirmed their predicted trypanocidal effects. Conclusions: We have built portable meta-classifiers capable of identifying small molecules with trypanocidal activity against amastigotes, the clinically most relevant stage of T. cruzi. The predictive ability of this meta-classifier was experimentally validated. Full article

Research on in vitro skin models has advanced remarkably, driven by a better understanding of the skin and the search for more ethical and efficient methods. The development of these models was initially motivated by the need for reduced animal testing and a faster and more ethical approach for the safety evaluation of cosmetic and pharmaceutical products. Stricter regulations and growing ethical awareness have driven further evolution, resulting in more refined and reliable methods. Diversity of cell types is crucial to replicating the complexity of human skin, including epithelial, dendritic, endothelial, and adipose cells, providing environments that closely mimic the physiological skin environment. This allows for more precise studies on skin interactions with cosmetic, dermatological, and pharmaceutical products. In vitro skin models have applications in toxicity testing, dermatological product evaluation, skin ageing studies, and drug research, reducing dependence on animal testing. This review presents a look at the different types of in vitro skin models developed for various applications, with a brief look at their strengths and drawbacks. Models developed for disease-specific applications are also covered. Techniques such as bioprinting and organ-on-a-chip have revolutionised the manufacturing of these models. Challenges persist, such as the need to improve vascularisation and faithfully replicate skin architecture. The promising future of these models points to an exciting path forward for dermatological research and the cosmetic industry. This review addresses the history and regulations of skin models, explores various skin models, and highlights the most recent advances, outlining future perspectives and offering a comprehensive overview. Full article

Parasites of the Leishmania genus are globally distributed and cause various clinical presentations in animals and humans, collectively known as leishmaniasis. The genomes of Leishmania and other trypanosomatids exhibit remarkable plasticity, shaped by several distinctive genetic features. Although these features can hinder the application of next-generation DNA sequencing (NGS) technologies, NGS data have been successfully used to characterize the whole-genome diversity of circulating Leishmania strains. The results complement and are broadly aligned with previous findings obtained with more traditional methods, offering greater resolution when working with geographically closer strains. In this review, we summarize advances over the past two decades in characterizing the genome diversity of Leishmania parasites using NGS strategies. We also discuss the application of these strategies to elucidate other aspects relevant to the epidemiology of these parasites, including their population structure and mode of reproduction. The vast majority of the studies to date have focused on species within the L. donovani/infantum complex or the L. (Viannia) subgenus, highlighting the need to incorporate other relevant underrepresented species and regions from both the Old and New World. Full article

The mitochondrial DNA of trypanosomatid parasites consists of thousands of catenated minicircles and dozens of maxicircles that form a complex network structure, the kinetoplast (kDNA). Although kDNA replication and segregation during mitotic division are well studied, its inheritance during genetic exchange events remains unclear. In Trypanosoma brucei, hybrids inherit minicircles biparentally but retain maxicircles from a single parent. Although biparental inheritance of minicircles has been described in natural Trypanosoma cruzi hybrids, this process has not been explored in laboratory-generated hybrids of this parasite. In the present study, we analyzed kDNA inheritance in T. cruzi experimental hybrids using a comprehensive minicircle hypervariable region (mHVR) database and genome sequencing data. Our findings revealed biparental inheritance of minicircles, with hybrid lines retaining mHVRs from both parents for over 800 generations. In contrast, maxicircles were exclusively inherited from one parent. Unexpectedly, we observed an increase in kDNA content in hybrids, affecting both minicircles and maxicircles, and exhibiting instability over time. To explain these findings, we propose a Replicative Mixing (REMIX) model, where the hybrid inherits one kinetoplast from each parent and they are replicated allowing minicircle mixing. Instead maxicircle networks remain physically separated, leading to uniparental fixation after segregation in the first cell division of the hybrid. This model challenges previous assumptions regarding kDNA inheritance and provides a new framework for understanding kinetoplast dynamics in hybrid trypanosomes. Full article

Background: Chagas disease and leishmaniasis remain public health concerns. Despite the existence of approved medications for the treatment of these diseases, most patients discontinue treatment due to long drug regimens and/or the severe side effects of these drugs. This leads to treatment failure and potential future drug resistance. Therefore, the search for new molecules with trypanocidal activity, low cytotoxicity, and high selectivity is essential to address this challenge. Methods: In this work, three series (a, b, and c) of pyridine-2,5-dicarboxylate esters were synthesized using different β-keto-esters bearing naturally occurring fragments and 1,2,3-triazine-1-oxides via the inverse electron demand Diels–Alder (IEDDA) reaction. The structural elucidation of the compounds was performed using NMR (¹H and ¹³C) and HRMS, and the crystal structure of compound 6a was also obtained. Furthermore, a biological assay was performed for all synthesized and characterized compounds to determine their cytotoxicity against Trypanosoma cruzi, Leishmania mexicana, and the J774.2 macrophage cell line. Finally, the in silico determination of their pharmacokinetic and toxicological properties was performed using the SwissADME and ProTox 3.0 platforms. Results: Compounds 3a, 4a, 5a, 4b, and 8c had the highest anti-Trypanosoma cruzi activity against both strains (IC₅₀ ≤ 56.68 µM). Compounds 8b, 10a, 9b, and 12b had considerable leishmanicidal activity against Leishmania mexicana against both strains (IC₅₀ ≤ 161.53 µM). Furthermore, in silico prediction of ADMET properties suggest that these pyridine compounds possess good pharmacokinetic profile. The results are also consistent with low in vitro cytotoxicity and high selectivity. Conclusions: The synthesized pyridine-2,5-dicarboxylate esters have promising activity against Trypanosoma cruzi and Leishmania mexicana, with low cytotoxicity and good drug-like properties, suggesting that these compounds are potential candidates for further evaluation as new treatments for Chagas disease and leishmaniasis. Full article

The actin cytoskeleton plays a crucial role in fundamental eukaryotic processes such as morphogenesis, motility, endocytosis, intracellular trafficking, and cell division. However, our understanding of actin and its associated proteins in trypanosomatid parasites like Leishmania remains limited. Over the past two decades, considerable progress has been made in elucidating the structure and functions of Leishmania actin and its core regulators. Notably, these findings are primarily derived from studies of the insect-stage promastigote form, while the roles of the actin machinery during the disease-causing amastigote stage within mammalian hosts remain largely unexplored. This review consolidates the current knowledge of actin and its interactors in Leishmania promastigotes, highlighting their potential roles in parasite development and stage-specific differentiation. Additionally, it explores the potential of targeting the cytoskeletal system as a strategy for novel therapeutic interventions against Leishmaniasis. The review concludes by identifying critical knowledge gaps and proposing future research directions to better understand actin-driven pathogenesis in this important human parasite. Full article

Sterol biosynthesis is crucial for the function of biological membranes and an important target for anti-protozoan/anti-fungal drugs. In the trypanosomatid parasite Leishmania major, the deletion of sterol C14-demethylase (C14DM) results in hypersensitivity to heat, increased plasma membrane fluidity, profound mitochondrial dysfunctions, and reduced virulence in mice. In this study, we show that C14DM-null mutants are defective in their tolerance to membrane-disrupting agents and osmotic stress and their ability to form autophagosomes. In addition, C14DM-null mutants exhibit a heightened sensitivity to anti-trypanosomatid drugs including antimony, ethidium bromide, and pentamidine. The combination of itraconazole (a C14DM antagonist) and pentamidine synergistically inhibits the growth of Leishmania parasites. These findings reveal new insight into the roles of sterol synthesis in protozoan pathogens and highlight the potential of using drug combinations to achieve better treatment outcomes. Full article

Neglected Tropical Diseases (NTDs) encompass a range of disorders, including infectious diseases caused by viruses, bacteria, parasites, fungi, and toxins, mainly affecting underprivileged individuals in developing countries. Among the NTDs, those caused by parasites belonging to the Trypanosomatidae family are particularly impacting and require attention, since the lack of financial incentives has led to constraints on the development of novel drugs to tackle them effectively. To circumvent the minor advances in drug discovery in this area, academic research emerges as a crucial player, namely through the identification and validation of new drug targets, thereby contributing to the development of more efficient, safe, and less expensive therapies against Trypanosomatidae infections. Noteworthy, this is a matter of utmost urgency since these diseases are endemic in countries with low socioeconomic standards. This review provides a comprehensive understanding of the current paradigm of NTDs caused by parasites belonging to the Trypanosomatidae family, addressing the ongoing limitations and challenges associated to the current chemotherapy solutions for these diseases and discussing the opportunities unravelled by recent research that led to the identification of new biomolecular targets that are common to Trypanosomatidae parasites. Among these, the unique properties of Trypanothione Synthetase (TryS) and Trypanothione Reductase (TryR), two key protozoan enzymes that are essential for the survival of Trypanosoma and Leishmania parasites, will be emphasised. In addition to a critical analysis of the latest advances in the discovery of novel molecules capable of inhibiting TryS and TryR, the possibility of dual targeting through a combination of TryS and TryR inhibitors will be addressed Full article

Information on Culicoides transmitted parasites, especially trypanosomatids, infecting animals and insects, is scarce. Our goal was to clarify the seasonal patterns of both Culicoides and trypanosomatids detected in these insects and the relationships between Culicoides abundance and meteorological parameters. UV light traps were used to collect biting midges in four study sites in 2022–2023; collected Culicoides females were dissected and analyzed using microscopy and PCR-based methods. Out of 1631 parous Culicoides females belonging to 14 different species, 6.5% were found to be infected with trypanosomatids (5.0% with at least three Trypanosoma species and 1.5% with monoxenous parasites). The highest Culicoides abundance was detected in June. The prevalence of trypanosomatids in biting midges increased during the summer (5.3% in June, 8.8% in July, and 11.2% in August). Temperature was recorded to be a presiding environmental gradient structuring Culicoides species composition, while wind speed and precipitation explained little of the variation. Our results indicate that both avian and mammalian trypanosomes can be found in these insects, although further research is needed to better understand the development of these parasites in biting midges and Culicoides vectorial capacity. Full article

Cyclopes, the smallest of all known anteaters, has an insectivorous diet and is arboreal, rarely descending to the ground. There are scarce reports on diseases and pathogenic agents affecting this taxon. Hemopathogens are pathogenic agents that inhabit the blood of various vertebrate species. Protozoa such as Trypanosoma spp., Leishmania spp., Hepatozoon spp., and members of the order Piroplasmida, as well as hemoplasmas and Rickettsial bacteria of the genera Anaplasma and Ehrlichia, are among the most important in this group. The transmission of these pathogens generally occurs through arthropod vectors, which act as intermediate hosts. In addition, infections caused by hemopathogens can have adverse effects on host health, contributing to population declines in susceptible species. This study investigated infection by protozoa and hemotropic bacteria in blood samples from free-ranging silky anteaters from Brazil, Peru, and Colombia using molecular detection methods. Sixteen samples were obtained during expeditions conducted in these countries. DNA was extracted from blood samples, and PCR assays were performed to detect parasites from the order Piroplasmida, Hepatozoon spp., trypanosomatid agents including Leishmania spp., Trypanosoma evansi, T. cruzi, and T. vivax, as well as hemotropic bacteria of the genera Ehrlichia, Anaplasma, and Mycoplasma sp. Nucleotide sequencing was performed on positive samples. Of the total samples analyzed, 62.5% (10/16) tested positive for hemotropic Mycoplasma, 50% (8/16) for T. evansi, and 6.2% (1/16) for T. cruzi. There is a significant gap in knowledge regarding the diversity of hemopathogens affecting the genus Cyclopes, and future studies are needed to understand how these infections may impact the health of individuals. Full article

Trypanosoma cruzi GenBank^® M21331 encodes for Antigen 36 (Ag 36), which is a tandemly repeated T. cruzi antigen. GenBank M21331 has a gene sequence similarity to human immune genes IFN-α, IFN-β, and IFN-γ, as well as to human TRIM genes. A BLAST-p search revealed that T. cruzi GenBank M21331 had seven gene sequences homologous to microtubule-associated protein (MAP) genes with a 100% amino acid sequence identity. There are 36 genes in the T. cruzi genome with >94% identity to GenBank M21331, and these genes encode proteins ranging in size from 38 to 2011 amino acids in length, the largest containing 20, 25, and 30 repeats of the Ag 36 thirty-eight-amino-acid-sequence motif. The purpose of this study was to perform a genetic and molecular comparative analysis of T. cruzi GenBank M21331 to determine if this gene sequence is unique to the T. cruzi clade, present in the T. brucei clade, and/or exists in other trypanosomatids. There are seven homologous genes to GenBank M21331 in T. cruzi, but only one homolog found of this gene in T. brucei. The MAP genes in T. cruzi appear to have expanded at least eleven-fold in number compared to similar MAP genes in T. brucei. The DNA sequences and functions of these MAP genes in their respective species and clades will be discussed and are a fascinating area for further scientific study. Full article